Hyaline membrane disease, HMD, is a common disorder of premature infants and is related to diffuse atelectesis, hypoxia and resultant respiratory impairment. More particularly, HMD relates to the lack of vital pulmonary material necessary for reducing surface tension in the airways of the alveoli. As a result, the alveoli or terminal respiratory sacs of patients suffering from HMD normally collapse. And, because the surface tension at the gas-liquid interface in HMD patients is elevated, their alveoli or terminal respiratory sacs are very difficult to reinflate. Consequently, HMD may be associated with significant morbidity and mortality, especially in premature infants.
Present treatments of HMD focus on using high concentrations of oxygen, positive pressure and/or mechanical ventilation to maintain adequate oxygenation. These therapies are complicated by oxygen and pressure related injuries as well as injuries resulting from the need to mechanically access the airway via endotracheal tubes. More recent studies, however, have supported the use of replacement pulmonary surfactant material for therapy of HMD and other syndromes associated with the lack of pulmonary surfactant material. Such therapy has included heretofore the use of aerosolized or liquid synthetic phospholipid mixtures, natural pulmonary surfactant material and various preparations of surfactant material prepared from animal lung. Surface tension lowering ability of the naturally derived preparations is in general better than in the synthetic lipid preparations. Also, preliminary studies using modified bovine surfactants have been promising. Problems with human and animal pulmonary preparations, however, have occurred which include lack of variability, possible infection and immunologic risks. Obviously, when treating patients for any disease including HMD, it is imperative to include only those active substances necessary to minimize possible immunologic consequences of therapy to the patients. Unfortunately, because the natural pulmonary surfactant material and the preparations available heretofore are in crude form, they are less specific and associated with possibly greater immunologic risks.
Natural pulmonary surfactant material is a complex material composed primarily of phospholipids and surfactant-associated proteins or apolipoproteins. The phospholipids, mainly phosphatidylcholine (PC), disaturated phosphatidylcholine (DSPC) and phosphatidylglycerol (PG), are of paramount importance for the physiological role of natural pulmonary surfactant material in reducing surface tension in the alveoli. Phospholipids, of which DSPC is the principal component, are synthesized in the endoplasmic reticulum of Type II epithelial cells, packaged into lamellar bodies, then secreted into the alveolar space by an exocytotic process. Several of the phospholipids are apparently not catabolized and resynthesized, but rather it is presently believed that they are reutilized primarily as intact molecules and constitute the major components of the naturally existing pulmonary surfactant material.
With respect to the surfactant-associated proteins or apolipoproteins, there is considerable disagreement as to their identity and utility. Nonetheless, there is increasing agreement among those with medical expertise in this area that in addition to the lung surfactant phospholipids, at least some of these apolipoproteins are vital for the full biological activity of the natural pulmonary surfactant material in reducing surface tension in the alveoli.
Surfactant-associated proteins or apolipoproteins include both serum and lung specific proteins. The major lung specific surfactant-associated protein of 30-40,000 daltons identified in lung surfactant by King et al, Isolation of Apoproteins from Canine Surfactant Material, Am J Physiol 244:788-795, 1973, is a glycoprotein rich in glycine and containing collagen-like regions rich in hydroxyproline. This protein, herein called SAP-35, is synthesized from 28-30,000 dalton translation products which undergo glycosylation, hydroxylation of proline residues and sulfhydryl-dependent cross-linking to form large oligomers which can be detected in the airway. Proteolytic fragments of SAP-35 have been identified in protein preparations isolated from lavage of patients with alveolar proteinosis and from other mammalian surfactants migrating as proteins of small molecular weight by Whitsett et al, Characteristics of Human Surfactant-Associated Glycoprotein(s) A, Pediatr Res 19:501-508, 1985. While the glycoprotein SAP-35 binds phospholipids and may confer the structural organization of tubular myelin to surfactant lipids, it remains unclear whether SAP-35 is required for the biophysical activity of surfactants. See King et al, Metabolism of the Apoproteins in Pulmonary Surfactant, J Appl Physiol 42:483-491, 1977.
Smaller lung specific surfactant-associated proteins have also been identified from a variety of mammalian surfactants. King et al, Am J Physiol 223:715-726, 1972, previously described a 10,000-12,000 dalton protein in pulmonary surfactant material; however, the origin of this protein or its distinction from others was not clarified. This protein described by King et al is now believed to be a fragment of the major glycoprotein SAP-35. Smaller surfactant-associated proteins, other than that reported by King et al, have been identified in alveolar lavage material from a number of species and with molecular weights of approximately 10,000 daltons in dog and rabbit, 10,500-14,000 daltons in rat, 11,500-16,500 daltons in pig, and 10,000 daltons in cow.
The nature and relationships among these various surfactant-associated proteins (SAPs) and the larger protein, SAP-35 or its fragments, have not been established. Nevertheless, the work of Suzuki et al, J Lipid Res 23:53-61, 1982, suggested that a small 15,000 daltons protein in pig alveolar lavage had a greater affinity for lipid than SAP-35. Suzuki et al unfortunately did not distinguish the proteins from the SAP-35 or its fragments or demonstrate if there exists surfactant properties in a purified state. Rather, Suzuki et al only suggested that this 15,000 dalton protein is possibly a physiological regulator for the clearance of alveolar phospholipid. Claypool et al, J Clin Invest 74:677-684, 1984, suggested that a small unidentified protein, isolated from rat alveolar lavage, increased the uptake of liposomes by cultured Type II epithelial cells. Work by Wang et al, Amino Acid Composition of Low Molecular Weight Hydrophobic Surfactant Apoproteins, Fed Proc 44:1024 (abstract), 1985, described two distinct small molecular weight proteins in surfactant from rat that are ethanol soluble. But like Suzuki et al, Wang et al failed to purify, identify activity, or characterize the protein to homogeneity. Wang et al instead suggested that these small molecular weight proteins may be involved with surfactant recycling. It also has been suggested that the smaller molecular weight proteins, such as those discussed above, possibly arise as proteolytic fragments of the larger SAP-35 molecule At present, however, it is unclear whether SAP-35, one or more of the smaller proteins, or all proteins together are active components imparting biophysical activity to natural mammalian pulmonary surfactant material.
In view of the present state of the art, there obviously are needs to clarify the nature and role of the surfactant-associated lung specific proteins and to determine the most effective means for treating and preventing HMD and other syndromes associated with lack or insufficient amounts of pulmonary surfactant material to maximize HMD therapy and to eliminate the disadvantages associated with HMD therapy available heretofore.